[Impact of cyclin-dependent kinase inhibitor p27 on resistance of ovarian cancer multicellular spheroids to taxol].

OBJECTIVE: To examine the expression of the cyclin-dependent kinase inhibitor p27 in ovarian cancer multicellular spheroid (MCS) and explore the reversal effect of p27-antisense oligodeoxynucleotide (p27-ASON) on taxol resistance in ovarian cancer cell lines.

METHODS: Three-dimensional culture was used to form MCS of human ovarian cancer cell lines A2780 and CAOV3. The MCSs were divided into 3 groups: MCSs transduced with p27-ASON or p27-sense oligodeoxynucleotide (p27-SON) by LipofectAMINE respectively and MCS without transduction. Monolayer A2780 and CAOV3 cells were cultured as controls. The cells were exposed to taxol of different concentrations (0.2, 2.0, 10.0, and 20.0 micro mol/L) for 24 hours. The expression of p27 in those cells was detected with Western blot and flow cytometry (FACS), and the subcellular distribution of p27 was detected by laser confocal microscopy before and after the experiment. The cell cycle profile and apoptosis were analyzed by FACS. The resistance to taxol was detected with trypan blue exclusion testing.

RESULTS: Compared with that in monolayer cells, expression of p27 in MCS cells was significantly higher (P(A2780) = 0.011, P(CAOV3) = 0.024). The percentage of cells in G0-G1 phase was significantly higher in the MCS cells than in the monolayer cells, and the percentages of cells in S and G2-M phases were significantly lower in MCS cells than in monolayer cells. The apoptotic rate of the monolayer cells was significantly higher than those of MCS cells treated with taxol of the concentration of 20.0 micro mol/L (P(A2780) = 0.003, P(CAOV3) = 0.015). The apoptotic rate of p27-ASON/MCS was significantly higher than that of MCS (P(A2780) = 0.022, P(CAOV3) = 0.036). There was no significant difference of apoptotic rate between p27-SON/MCS and MCS (P(A2780) = 0.412, P(CAOV3) = 0.071). The monolayer A2780 and CAOV3 cells formed compact spheroids after three-dimensional culture. Cells transduced with p27-SON formed compact MCS too. However, cells tranduced with p27-ASON formed loose collections of cells easy to shatter. The expression of p27 in p27-ASON/MCSs was downregulated.

CONCLUSION: The drug resistance of ovarian cancer MCS is related to upregulation of p27. P27-ASON reverses the resistance of ovarian cancer to taxol, thus increasing the chemotherapeutic sensitivity of ovarian cancer cells.